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1.
On the molecular weight of mitochondrially synthesized subunits of rat liver cytochrome oxidase. 总被引:2,自引:0,他引:2
The subunit composition of cytochrome c oxidase from rat liver mitochondria was studied by dodecylsulfate polyacrylamide gel electrophoresis. The apparent molecular weight of the seven subunits are in reasonable agreement with published data on cytochrome c oxidase subunits from other sources. Two additional subunits were found if the electrophoresis was performed with 8m urea, due to splitting of the smallest subunit. Performic acid oxidation of the isolated subunits I and II increased the apparent molecular weights from 38000 to 48000 and from 24500 to 29000, respectively, accompained by a normalization of the anomalous behaviour of subunit I in the Ferguson plot. It is suggested that performic acid, by splitting extremely inaccessible disulfide bridges, mediates full complexing of the subunits by dodecylsulfate, thus permitting the determination of the real molecular weights by dodecylsulfate polyacrylamide gel electrophoresis. 相似文献
2.
R Michel A Liebl W Machleidt J Otto W Neupert 《Hoppe-Seyler's Zeitschrift für physiologische Chemie》1975,356(10):1595-1604
A polypeptide with a molecular weight of 8 500 (HP 8 500) was isolated from the mitochondrial membrane of the nuclear mutant cni-1 of Neurospora crassa. This mutant is characterized by a cyanide-insensitive respiration and by a deficiency in the cytochromes aa3 and b. The polypeptide is synthesized on mitochondrial ribosomes. It has an extremely hydrophobic character; it is insoluble in aqueous media in the absence of sodium dodecylsulfate and is soluble in acid chloroform/methanol. It lacks histidine. The polar amino acids lysine, arginine, aspartic acid, glutamic acid, serine and threonine make up only 25% of the total amino acids on a mole-percent basis. The N-terminal amino acid is tyrosine. The possible function of this polypeptide in the mitochondrial membrane is discussed. 相似文献
3.
Three groups of proteins can be clearly discriminated in the total protein of L cell polysomes by selective labelling in the presence of low doses of actinomycin D and two-dimensional polyacrylamide/dodecylsulfate gel electrophoresis followed by autoradiography: (a) structural ribosomal proteins which are not labelled in the presence of actinomycin D and form stained non-radioactive spot in gels; (b) exchangeable ribosomal proteins which are labelled in the presence of actinomycin D and stained radioactive spots; (c) non-ribosomal proteins which are detectable only by autoradiography of gels. The large and small subunits of L cell ribosomes contain respectively 45 and 34 ribosomal proteins with molecular weights less than or equal to 50 000; seven of the large subunit proteins and nine of the small subunit proteins are exchangeable. Most of the non-ribosomal proteins migrate in the region of the related to the separation of the ribosomal proteins of mammalian cells and the possible significance of the presence of non-ribosomal proteins in polysomes are discussed. 相似文献
4.
5.
R Michel A Liebl A Hartmann W Neupert 《Hoppe-Seyler's Zeitschrift für physiologische Chemie》1976,357(3):415-426
Gel electrophoretic analysis of mitochondrial membranes from Neurospora crassa shows the presence of a polypeptide fraction with apparent molecular weights of 7000 - 1200, which is synthesized on mitochondrial ribosomes. This fraction comprises between 10 and 50% of total mitochondrial translation products. Evidence is presented that the major part of this fraction is derived from components with higher apparent molecular weights by proteolytic activity. The proteolytic activity is located in vesicles which are co-isolated with mitochondria upon differential centrifugation. The activity is strongly enhanced by application of detergents such as sodium dodecylsulfate and Triton. Proteins synthesized on mitochondrial as well as cytoplasmic ribosomes are subject to proteolytic breakdown. This proteolysis can be blocked by addition of inhibitors such as diisopropylfluorphosphate to isolated mitochondria. Similar observations were made with Schizosaccharomyces pombe. In Neurospora, the amount of mitochondrial translation products with apparent molecular weights of less than 12000 is low in mitochondria from cells treated with cycloheximide for 1 h and high in mitochondria from cells treated with cycloheximide for 5 min. This observation is explained by the finding that proteinase activity in mitochondrial preparations decreases exponentially with a t1/2 of 20 min during preincubation of cells with cycloheximide. Procedures are described to remove or block contaminating proteinase activity. The results appear to be relevant for the interpretation of many data obtained from experiments in which this puzzling kind of artifact has not been sufficiently considered. 相似文献
6.
O'Brien TW 《IUBMB life》2003,55(9):505-513
Mammalian mitochondrial ribosomes (55S) differ unexpectedly from bacterial (70S) and cytoplasmic ribosomes (80S), as well as other kinds of mitochondrial ribosomes. Typical of mammalian mitochondrial ribosomes, the bovine mitochondrial ribosome has been developed as a model system for the study of human mitochondrial ribosomes, to address several questions related to the structure, function, biosynthesis and evolution of these interesting ribosomes. Bovine mitochondrial ribosomal proteins (MRPs) from each subunit have been identified and characterized with respect to individuality and electrophoretic properties, amino acid sequence, topographic disposition, RNA binding properties, evolutionary relationships and reaction with affinity probes of ribosomal functional domains. Several distinctive properties of these ribosomes are being elucidated, including their antibiotic susceptibility and composition. Human mitochondrial ribosomes lack several of the major RNA stem structures of bacterial ribosomes but they contain a correspondingly higher protein content (as many as 80 proteins), suggesting a model where proteins have replaced RNA structural elements during the evolution of these ribosomes. Despite their lower RNA content they are physically larger than bacterial ribosomes, because of the 'extra' proteins they contain. The extra proteins in mitochondrial ribosomes are 'new' in the sense that they are not homologous to proteins in bacterial or cytoplasmic ribosomes. Some of the new proteins appear to be bifunctional. All of the mammalian MRPs are encoded in nuclear genes (a separate set from those encoding cytoplasmic ribosomal proteins) which are evolving more rapidly than those encoding cytoplasmic ribosomal proteins. The MRPs are imported into mitochondria where they assemble coordinately with mitochondrially transcribed rRNAs into ribosomes that are responsible for translating the 13 mRNAs for essential proteins of the oxidative phosphorylation system. 相似文献
7.
Earlier studies have shown that cytochrome c oxidase from bakers' yeast is an oligomeric enzyme which contains three polypeptides (I to III) synthesized on mitochondrial ribosomes and four polypeptides (IV to VII) synthesized on cytoplasmic ribosomes. These polypeptide subunits have now been isolated by a simple protocol which utilizes differences in polypeptide charge, solubility, and size. Their molecular weights determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, gel filtration in the presence of guanidine hydrochloride, and amino acid analysis were: I, 40,000; II, 33,000; III, 22,000; IV, 14,500; V, 12,700; VI, 12,700; and VII, 4,600. All seven polypeptide subunits exhibited acidic isoelectric points; cytoplasmically made subunits were more acidic than mitochondrially made ones. The amino acid composition of two mitochondrially made subunits and two cytoplasmically made subunits was determined. The two mitochondrial translation products, I and II, contained only 34.7% and 42.1% polar amino acids, respectively, whereas the two cytoplasmic translation products, IV and VI, contained 48.3% and 49.3%, respectively. This agreed with the observation that Subunits I and II are very insoluble, requiring detergents for solubility, whereas Subunits IV and VI are water-soluble in the absence of any added detergent. These results indicate that the cytochrome c oxidase subunits synthesized on mitochondrial and cytoplasmic ribosomes are fundamentally different in size, isoelectric properties, and hydrophobicity. They also suggest the possibility that at least some of the mitochondrially made subunits are buried in the lipid phase of the mitochondrial inner membrane. 相似文献
8.
The proteins were extracted from purified 40-S ribosomes derived from wheat germ and Artemia salina and separated by carboxymethylcellulose ion-exchange chromatography. Approximately four proteins from Artemia and four proteins from wheat germ were separated in a state of high purity. All proteins were identified by co-electrophoresis using a two-dimensional polyacrylamide gel system. A total of 30 unique proteins were found for Artemia and 32 proteins for wheat. The molecular weights of all proteins were estimated by sodium dodecylsulfate gel electrophoresis. Assuming each protein to be present in one copy per 40-S ribosome, the total protein molecular weight was estimated to be 560,000 associated with Artemia 40-S particles and 550,000 associated with wheat germ 40-S ribosomes. 相似文献
9.
G Huynh-Van-TanSchapira 《European journal of biochemistry》1978,85(1):271-281
Free cytoplasmic messenger ribonucleoproteins have been prepared with a high yield in a zonal rotor. They are rapidly labelled by [3H]uridine. Their sedimentation coefficients are 14.6 S and 19.4 S. Their protein contents are higher than in ribosomes, as indicated by a shift towards greater wavelength in their ultraviolet spectra and a buoyant density of 1.39 g cm-2 in CsCl. They do not cross-react with antiribosome antibodies and their protein patterns in sodium dodecylsulfate and two-dimensional polyacrylamide gel electrophoresis are different from ribosomes'. They may contain up to 30 proteins. The proteins are acidic and their molecular weights range from 22 000 to 100 000. When compared to chromatin they behave more like non-histone proteins than histones. 相似文献
10.
About ScienceDirect 《Biochimica et Biophysica Acta (BBA)/General Subjects》1982,719(3):453-456
The photoaffinity ligand 8-azidoadenosine 3′,5-monophosphate was employed to label cAMP binding proteins in both fractionated and freeze-thawed rabbit gastric glands. Fractionated glands incorporated the azido-cAMP label primarily into two cytosolic proteins with apparent molecular weights of 58 000 and 48 000. No enrichment of label was found in fractions containing basolateral or apical membranes. DEAE-cellulose chromatography of the cytosolic proteins resulted in the separation of two cAMP-dependent protein kinase peaks. Azido cAMP labelling of each peak suggested the initial peak contained type I cAMP-dependent protein kinase while the second peak contained the type II kinase. Labelling of ‘resting’ gastric glands resulted in radioactive proteins of apparent molecular weights of 58 000.and 48 000. When gastric glands were stimulated to produce acid by the addition of 10?4 M histamine or 1 mM dibutyryl cAMP there was 32–44% dimunition of ligand incorporation compared to control glands. The results strongly suggest that histamine- mediated stimulus-secretion coupling in gastric glands involves activation of parietal cell cAMP-dependent protein kinases. 相似文献
11.
The photoaffinity ligand 8-azidoadenosine 3',5-monophosphate was employed to label cAMP binding proteins in both fractionated and freeze-thawed rabbit gastric glands. Fractionated glands incorporated the azido-cAMP label primarily into two cytosolic proteins with apparent molecular weights of 58 000 and 48 000. No enrichment of label was found in fractions containing basolateral or apical membranes. DEAE-cellulose chromatography of the cytosolic proteins resulted in the separation of two cAMP-dependent protein kinase peaks. Azido cAMP labelling of each peak suggested the initial peak contained type I cAMP-dependent protein kinase while the second peak contained the type II kinase. Labelling of 'resting' gastric glands resulted in radioactive proteins of apparent molecular weights of 58 000 and 48 000. When gastric glands were stimulated to produce acid by the addition of 10(-4) M histamine or 1 mM dibutyryl cAMP there was 32-44% dimunition of ligand incorporation compared to control glands. The results strongly suggest that histamine-mediated stimulus-secretion coupling in gastric glands involves activation of parietal cell cAMP-dependent protein kinases. 相似文献
12.
Protein composition of mitochondrial ribosomes of the yeast Saccharomyces cerevisiae was analysed by two-dimensional electrophoresis. The small (37S) mitoribosomal subunit contains 36 different polypeptides with molecular weights ranging from 10,000 to 60,000. The large (50S) subunit is composed of 41 proteins with molecular weights from 10,000 to 43,000. The molecular weights of mitoribosomal small and large subunits are 1.85 MDa and 2.35 MDa, respectively. Proteins represent 60-62% and 42-45% of the total mass of 37S and 50S subunits respectively. On the basis of the protein content and molecular weights of individual proteins we conclude that all mitoribosomal proteins are present in the mitoribosome in equimolar proportions. 相似文献
13.
O'Brien TW 《Gene》2002,286(1):73-79
Mitochondrial ribosomes comprise the most diverse group of ribosomes known. The mammalian mitochondrial ribosomes (55S) differ unexpectedly from bacterial (70S) and cytoplasmic ribosomes (80S), as well as other kinds of mitochondrial ribosomes. The bovine mitochondrial ribosome has been developed as a model system for the study of human mitochondrial ribosomes to address several questions related to the structure, function, biosynthesis and evolution of these interesting ribosomes. Bovine mitochondrial ribosomal proteins (MRPs) from each subunit have been identified and characterized with respect to individuality and electrophoretic properties, amino acid sequence, topographic disposition, RNA binding properties, evolutionary relationships and reaction with affinity probes of ribosomal functional domains. Several distinctive properties of these ribosomes are being elucidated, including their antibiotic susceptibility and composition. Mammalian mitochondrial ribosomes lack several of the major RNA stem structures of bacterial ribosomes but they contain a correspondingly higher protein content (as many as 80 proteins), suggesting a model where proteins have replaced RNA structural elements during the evolution of these ribosomes. Despite their lower RNA content they are physically larger than bacterial ribosomes, because of the 'extra' proteins they contain. The extra proteins in mitochondrial ribosomes are 'new' in the sense that they are not homologous to proteins in bacterial or cytoplasmic ribosomes. Some of the new proteins appear to be bifunctional. All of the mammalian MRPs are encoded in nuclear genes (a separate set from those encoding cytoplasmic ribosomal proteins) which are evolving more rapidly than those encoding cytoplasmic ribosomal proteins. The MRPs are imported into mitochondria where they assemble coordinately with mitochondrially transcribed rRNAs into ribosomes that are responsible for translating the 13 mRNAs for essential proteins of the oxidative phosphorylation system. Interest is growing in the structure, organization, chromosomal location and expression of genes for human MRPs. Proteins which are essential for mitoribosome function are candidates for involvement in human genetic disease. 相似文献
14.
Two sPPases (soluble inorganic pyrophosphatases, EC 3.6.1.1) have been isolated from the microalga Chlamydomonas reinhardtii. Both are monomeric proteins of organellar localization, the chloroplastic sPPase I [Cr (Ch. reinhardtii)-sPPase I, 30 kDa] is a major isoform and slightly larger protein than the mitochondrial sPPase II (Cr-sPPase II, 24 kDa). They are members of sPPase family I and are encoded by two different cDNAs, as demonstrated by peptide mass fingerprint analysis. Molecular phylogenetic analyses indicated that Cr-sPPase I is closely related to other eukaryotic sPPases, whereas Cr-sPPase II resembles its prokaryotic counterparts. Chloroplastic sPPase I may have replaced a cyanobacterial ancestor very early during plastid evolution. Cr-sPPase II orthologues are found in members of the green photosynthetic lineage, but not in animals or fungi. These two sPPases from photosynthetic eukaryotes are novel monomeric family I sPPases with different molecular phylogenies and cellular localizations. 相似文献
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16.
Ribosome binding to the Oxa1 complex facilitates co-translational protein insertion in mitochondria 总被引:12,自引:0,他引:12
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The Oxa1 translocase of the mitochondrial inner membrane facilitates the insertion of both mitochondrially and nuclear-encoded proteins from the matrix into the inner membrane. Most mitochondrially encoded proteins are hydrophobic membrane proteins which are integrated into the lipid bilayer during their synthesis on mitochondrial ribosomes. The molecular mechanism of this co-translational insertion process is unknown. Here we show that the matrix-exposed C-terminus of Oxa1 forms an alpha-helical domain that has the ability to bind to mitochondrial ribosomes. Deletion of this Oxa1 domain strongly diminished the efficiency of membrane insertion of subunit 2 of cytochrome oxidase, a mitochondrially encoded substrate of the Oxa1 translocase. This suggests that co-translational membrane insertion of mitochondrial translation products is facilitated by a physical interaction of translation complexes with the membrane-bound translocase. 相似文献
17.
Proteomic studies have demonstrated that yeast mitochondria contain roughly 1000 different proteins. Only eight of these proteins are encoded by the mitochondrial genome and are synthesized on mitochondrial ribosomes. The remaining 99% of mitochondrial precursors are encoded within the nuclear genome and after their synthesis on cytosolic ribosomes must be imported into the organelle. Targeting of these proteins to mitochondria and their import into one of the four mitochondrial subcompartments--outer membrane, intermembrane space (IMS), inner membrane and matrix--requires various membrane-embedded protein translocases, as well as numerous chaperones and cochaperones in the aqueous compartments. During the last years, several novel protein components involved in the import and assembly of mitochondrial proteins have been identified. The picture that emerges from these exciting new findings is that of highly dynamic import machineries, rather than of regulated, but static protein complexes. In this review, we will give an overview on the recent progress in our understanding of mitochondrial protein import. We will focus on the presequence translocase of the inner mitochondrial membrane, the TIM23 complex and the presequence translocase-associated motor, the PAM complex. These two molecular machineries mediate the multistep import of preproteins with cleavable N-terminal signal sequences into the matrix or inner membrane of mitochondria. 相似文献
18.
Cyclic AMP receptor protein from yeast mitochondria: submitochondrial localization and preliminary characterization. 总被引:1,自引:0,他引:1
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We have identified and characterized a cyclic AMP receptor protein in mitochondria of the yeast Saccharomyces cerevisiae. The binding is specific for cyclic nucleotides, particularly for cyclic AMP which is bound with high affinity (Kd of 10(-9) M) at 1 to 5 pmol/mg of mitochondrial protein. The mitochondrial cyclic AMP receptor is synthesized on cytoplasmic ribosomes and has an apparent molecular weight of 45,000 as determined by photoaffinity labeling. It is localized in the inner mitochondrial membrane and faces the intermembrane space. Cross-contamination of mitochondrial inner membranes by plasma membranes or soluble cytoplasmic proteins is excluded. 相似文献
19.
The mechanisms of interaction of nuclear and mitochondrial genes in biogenesis of mitochondria are discussed in this review. Brief characterization of yeast mitochondrial genes and their products is presented. The mechanism of nuclear and mitochondrial control of expression of the mosaic genes in mitochondria is described. The data on the processing of imported mitochondrial proteins synthesized on cytoplasmic ribosomes are presented. The possibility of existence of common proteins encoded for by common genes and possessing similar functions in the cytoplasm and mitochondria is discussed. A hypothesis is put forward considering the role of common proteins in coordination of nuclear and mitochondrial genes' expression in biogenesis of mitochondria. 相似文献
20.
F Paltauf 《European journal of biochemistry》1978,85(1):263-270
The terminal step during aerobic plasmalogen biosynthesis is catalyzed by a microsomal desaturase system which converts 1-O-alkyl-2-acyl-sn-glycerophosphoethanolamine to 1-O-alk-1'-enyl-2-acyl-sn-glycerophosphoethanolamine (ethanolamine plasmalogen). The reaction depends on oxygen and NAD(P)H and is stimulated 3-10-fold by soluble activating factors contained in the 100 000 X g supernatant. Two stimulating proteins have been isolated from pig kidney; the partially purified proteins have identical molecular weights (27 000) but differ in their respective isoelectric points (protein I, 5.1 and protein II, 4.9). Both proteins behave identically in the biochemical studies conducted. Exogenous substrate binds to the stimulating proteins; the transfer of ethanolamine, but not of choline phospholipids, from liposomes to microsomes is enhanced by the stimulating proteins. They stimulate plasmalogen synthesis from either exogenous or endogenous substrate (synthesized from alkylglycerophosphoethanolamine by microsomal transacylases). The stimulating proteins have no enzymatic activity themselves; it is suggested that they affect events within the membrane and function as specific mediators between the membrane-bound enzyme system and the lipophilic substrate. 相似文献